1. Field of the Invention
The present invention relates generally to the field of immunoconjugates and, more particularly, to the use of immunoconjugates in the diagnosis and treatment of cancer. The invention also relates to the treatment of breast and cervical carcinoma with cytotoxic conjugates of monoclonal antibodies (MoAbs) and gelonin, a ribosomal inhibiting protein.
2. Background of the Invention
Breast cancer and cervical cancer are two of the leading causes of death from malignancy in women in the Western world. Surgical removal of localized malignancies has proven effective only when the disease has not spread beyond the primary lesion. Once the disease has spread, the surgical procedures must be supplemented with other more general procedures to eradicate the diseased or malignant cells. Most of the commonly utilized supplementary procedures such as irradiation or chemotherapy are not localized to the tumor cells and, although they have a proportionally greater destructive effect on malignant cells, often affect normal cells to some extent.
Many tumors express antigens or antigenic determinants which are either expressed very weakly or not expressed at all by normal cells. Some tumor cells express antigens which are expressed by embryonic cell types but are not expressed by normal cells of a mature animal. These abnormally expressed antigens are known as tumor-associated antigens. The antigens expressed by tumors are specific in that while a particular antigen may be expressed by more than one tumor, it is usually expressed by all or most cells of the particular tumors which express it. A tumor cell may express one or more than one tumor-associated antigen. These tumor-associated antigens may be expressed on the surface of the cell (cell surface antigen), may be secreted by the tumor cell (secreted antigens) or may remain inside the cell (intercellular antigen).
The presence of these tumor-associated antigens has been utilized to detect, diagnose and localize the tumor. In some cases the presence of the tumor-associated antigens on the tumor cells has allowed the targeting of specific drugs and treatment means specifically to the tumor cells.
Antibodies are proteins normally produced by the immune system of an animal in response to foreign antigens or antigenic determinants. Antibodies bind to the specific antigen to which they are directed. Monoclonal antibodies directed to specific antigens or antigenic determinants may be prepared in large quantities.
One method of targeting chemotherapeutic agents to tumor cells and to diminishing their effects on normal cells has been made possible with the development of MoAbs directed against antigens on the tumor cells which do not occur on normal cells.
Antibodies may be labelled in order to allow their use for localization and treatment of malignant diseases. Antibodies, coupled to drugs, may be used as a delivery system by which the drug is targeted to a specific tumor cell type against which the antibody is directed. Antibodies may also be coupled to toxins and thus act as a delivery system to target the toxins directly to specific tumor cells.
Gelonin is a glycoprotein (M.W. 29-30,000) purified from the seeds of Gelonium multiforum. Gelonin belongs to a class of potent ribosomal inactivating plant toxins. Other members of this class of ribosomal inactivating plant toxins are the chains of Abrin, Ricin and Modeccin. Gelonin, like abrin and ricin, inhibits protein synthesis by damaging the 60S sub-unit of mammalian ribosomes. The inactivation of ribosomes is irreversible, does not appear to involve co-factors and occurs with an efficiency which suggests that Gelonin acts enzymatically.
Numerous prior workers have suggested or reported linking cytotoxic agents to antibodies to make xe2x80x9cimmunotoxins.xe2x80x9d Of particular interest have been immunotoxins of monoclonal antibodies conjugated to the enzymatically active portions (A chains) of toxins of bacterial or plant origin such as Ricin or Abrin. Nevelle and York, Immunol. Rev. (1982) 62: 75-91; Ross et al., European J. Biochem. (1980) 104; Vitteta et al., Immunol. Rev. (1982) 62: 158-183; Ross et al., Cancer Res. (1982) 42: 457-464; Trowbridge and Domingo Nature (Cond.) (1981) 294: 171-173.
Gelonin and ricin are among the most active toxins in inhibiting protein synthesis on a protein weight basis. Gelonin is 10 to 1000 times more active in inhibiting protein synthesis than the ricin A chain. Peptides like ricin and abrin are composed of two chains, an A chain which is the toxic unit and a B chain which acts by binding to cells. Unlike ricin and abrin, gelonin is composed of a single chain, and, lacking a B chain for binding to cells, is itself non-toxic to intact cells. (Stirpe, et al. J. Biol. Chem. 255: 6947-6953 (1980)). Mammalian cells apparently lack the ability to bind and/or to internalize the native gelonin molecule. Conjugates of gelonin to a tumor-targeting monoclonal antibody such as the monoclonal antibody 15A8 directed to an antigen present on certain tumor cells such as breast cancer cells, provides both a specific method for binding the gelonin to the cell and a route for internalization of the gelonin-antibody complex. Among the advantages of using the toxin gelonin over using toxins such as ricin A chain is its reduced toxicity to normal tissues compared to ricin A chain. Gelonin coupled to and anti-tumor associated antigen monoclonal antibody is an active and selective immunotoxic agent for tumor therapy.
Since the antibody to which the drug, toxin or radioactive label is coupled binds only to tumor cells expressing a specific antigen, only the tumor cells are killed. However, radiation from the radiolabeled compounds is not limited solely to the tumor cells in which the radiation is taken up. Radiolabelled antibodies suffer from problems which limit or complicate their use as the therapeutic agents. For example, metabolic or enzymatic degradation of the antibody may release the radiolabel and allow it to distribute to other tissues such as kidneys or bone marrow, causing unacceptable radiation damage to these organs.
The present invention provides immuno conjugates of an antibody which recognizes the 15A8 antigen on breast cancer and cervical cancer cells. In one embodiment the antibody is coupled with a toxin selected from the group consisting of gelonin, ricin A chain and abrin A chain. In another embodiment the 15A8 antibody may be coupled with a cytocidal drug such as adriamycin or a biological response modifier such as a lymphokine or cytokine. In another embodiment the antibody may be labeled with a detectable label such as a radiolabel, a chemiluminescer, a fluorescer, or an enzyme label. The cytocidal immunoconjugates are useful to treat and prevent recurrence of tumor-associated 15A8-bearing tumors by administration of these cytocidal immunoconjugates to an individual in need of such treatment. The detectably labeled 15A8 immunoconjugates are useful for diagnosis and localization of tumors by techniques known to those in the art. These labeled immunoconjugates are also useful to assay for the presence of the 15A8 antigen in biological specimens and for localizing the tumor site in vivo by means known to those of skill in the art.
One of the objects of the present invention is to provide a cytotoxic composition which would specifically bind to and kill tumor cells. Specifically, it is an object of the present invention to provide a cytotoxic composition which would specifically bind to and kill tumor cells which express the 15A8 antigen (the antigen recognized by the monoclonal antibodies disclosed and claimed in U.S. patent application Ser. No. 29,373 filed Mar. 23, 1987). Another aspect of the invention concerns a method of killing human breast cancer cells, cervical carcenoma cells, or other tumor cells expressing the 15A8 antigen by contacting the cells with a cytocidally effective amount of an immunotoxin.
It is a further object of the present invention that such a composition would be toxic to tumor cells and would cause minimal injury to normal tissue.
It is a further object of the present invention to provide a composition comprising an antibody directed to a 22 kD 15A8 tumor-associated antigen. It was a further object of the present invention to provide a labelled antibody useful in the detection, diagnosis and localization of tumors expressing the 15A8 antigen.